Aptamers to PDGF and VEGF and their use in treating PDGF and VEGF mediated conditions

Abstract

Aptamers that bind PDGF and aptamers that bind VEGF are provided. In addition, aptamer constructs comprising a PDGF aptamer and a VEGF aptamer are provided. Pharmaceutical compositions comprising the aptamers and aptamer constructs are provided, as well as methods of treating conditions using the aptamers and aptamer constructs.

What is claimed is: 
     
       1. An aptamer comprising the sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 512) 
                 
                     
                   5′-ACAL n ZGZAZGL m ZLZ-3′ 
                 
             
                
                
               
            
           
         
         wherein; 
         each Z is, independently, a modified pyrimidine; 
         each L is independently selected from a substituted or unsubstituted C 2 -C 50  linker, a polyethylene glycol linker, and a modified or unmodified nucleotide; 
         at least one, at least two, at least three, at least four, or at least five nucleosides comprise a 2′-OMe; 
         n is 1 to 5; and 
         m is 1 to 10. 
       
     
     
       2. The aptamer of  claim 1 , wherein each Z is independently selected from 5-(N-benzylcarboxyamide)-2′-deoxyuridine (BndU), 5-(N-benzylcarboxyamide)-2′-O-methyluridine, 5-(N-benzylcarboxyamide)-2′-fluorouridine, 5-(N-phenethylcarboxyamide)-2′-deoxyuridine (PedU), 5-(N-thiophenylmethylcarboxyamide)-2′-deoxyuridine (ThdU), 5-(N-isobutylcarboxyamide)-2′-deoxyuridine (iBudU), 5-(N-isobutylcarboxyamide)-2′-O-methyluridine, 5-(N-isobutylcarboxyamide)-2′-fluorouridine, 5-(N-tryptaminocarboxyamide)-2′-deoxyuridine (TrpdU), 5-(N-tryptaminocarboxyamide)-2′-O-methyluridine, 5-(N-tryptaminocarboxyamide)-2′-fluorouridine, 5-(N-[1-(3-trimethylamonium)propyl]carboxyamide)-2′-deoxyuridine chloride, 5-(N-naphthylmethylcarboxyamide)-2′-deoxyuridine (NapdU), 5-(N-naphthylmethylcarboxyamide)-2′-O-methyluridine, 5-(N-naphthylmethylcarboxyamide)-2′-fluorouridine, and 5-(N-[1-(2,3-dihydroxypropyl)]carboxyamide)-2′-deoxyuridine). 
     
     
       3. The aptamer of  claim 1 , wherein n is 1, 2, 3, or 4; and wherein m is 1, 2, 3, 4, 5, 6, 7, 8, or 9. 
     
     
       4. The aptamer of  1  further comprising at least one, at least two, at least three, at least four, or at least five internucleoside linkages are phosphorothioate linkages. 
     
     
       5. The aptamer of  1 , wherein the aptamer binds PDGF with an affinity of less than 10 nM, less than 5 nM, less than 2 nM, or less than 1 nM. 
     
     
       6. The aptamer of  claim 1 , wherein the aptamer inhibits PDGF-mediated phosphorylation of a PDGF receptor. 
     
     
       7. The aptamer of  claim 1 , wherein L is independently, for each occurrence, selected from the group consisting of a substituted or unsubstituted C 2 -C 10  linker, a substituted or unsubstituted C 2 -C 8  linker, a substituted or unsubstituted C 2 -C 6  linker, a substituted or unsubstituted C 2 -C 5  linker, a substituted or unsubstituted C 2 -C 4  linker, a substituted or unsubstituted C 3  linker, a G, a C and an A. 
     
     
       8. A method comprising administering to a subject in need a therapeutically effective amount of the aptamer of  claim 1 , wherein the method treats and/or prevents a conditions selected from the group consisting of macular degeneration, an ophthalmic condition, fibrosis, a cardiovascular disease and a cancer. 
     
     
       9. An aptamer comprising the sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 512) 
                 
                     
                   5′-ACAL n ZGZAZGL m ZLZ-3′ 
                 
             
                
                
               
            
           
         
         wherein; 
         each Z is, independently, selected from the group consisting of a 5-(N-benzylcarboxyamide)-2′-deoxyuridine (BndU), 5-(N-benzylcarboxyamide)-2′-O-methyluridine, 5-(N-benzylcarboxyamide)-2′-fluorouridine, 5-(N-phenethylcarboxyamide)-2′-deoxyuridine (PedU), 5-(N-thiophenylmethylcarboxyamide)-2′-deoxyuridine (ThdU), 5-(N-isobutylcarboxyamide)-2′-deoxyuridine (iBudU), 5-(N-isobutylcarboxyamide)-2′-O-methyluridine, 5-(N-isobutylcarboxyamide)-2′-fluorouridine, 5-(N-tryptaminocarboxyamide)-2′-deoxyuridine (TrpdU), 5-(N-tryptaminocarboxyamide)-2′-O-methyluridine, 5-(N-tryptaminocarboxyamide)-2′-fluorouridine, 5-(N-[1-(3-trimethylamonium)propyl]carboxyamide)-2′-deoxyuridine chloride, 5-(N-naphthylmethylcarboxyamide)-2′-deoxyuridine (NapdU), 5-(N-naphthylmethylcarboxyamide)-2′-O-methyluridine, 5-(N-naphthylmethylcarboxyamide)-2′-fluorouridine, and 5-(N-[1-(2,3-dihydroxypropyl)]carboxyamide)-2′-deoxyuridine); 
         each L is independently selected from a substituted or unsubstituted C 2 -C 50  linker, a polyethylene glycol linker, and a modified or unmodified nucleotide; 
         n is 1 to 5; and 
         m is 1 to 10. 
       
     
     
       10. The aptamer of  claim 9  further comprising at least one, at least two, at least three, at least four, or at least five nucleosides comprise a 2′-OMe. 
     
     
       11. The aptamer of  claim 9 , wherein n is 1, 2, 3, or 4; and wherein m is 1, 2, 3, 4, 5, 6, 7, 8, or 9. 
     
     
       12. The aptamer of  9  further comprising at least one, at least two, at least three, at least four, or at least five internucleoside linkages are phosphorothioate linkages. 
     
     
       13. The aptamer of  9 , wherein the aptamer binds PDGF with an affinity of less than 10 nM, less than 5 nM, less than 2 nM, or less than 1 nM. 
     
     
       14. The aptamer of  claim 9 , wherein the aptamer inhibits PDGF-mediated phosphorylation of a PDGF receptor. 
     
     
       15. The aptamer of  claim 9 , wherein L is independently, for each occurrence, selected from the group consisting of a substituted or unsubstituted C 2 -C 10  linker, a substituted or unsubstituted C 2 -C 8  linker, a substituted or unsubstituted C 2 -C 6  linker, a substituted or unsubstituted C 2 -C 5  linker, a substituted or unsubstituted C 2 -C 4  linker, a substituted or unsubstituted C 3  linker, a G, a C and an A. 
     
     
       16. A method comprising administering to a subject in need a therapeutically effective amount of the aptamer of  claim 9 , wherein the method treats and/or prevents a conditions selected from the group consisting of macular degeneration, an ophthalmic condition, fibrosis, a cardiovascular disease and a cancer. 
     
     
       17. An aptamer comprising the sequence: 
       
         
           
                 
                 
               
                     
                   (SEQ ID NO. 512) 
                 
                     
                   5′-ACAL n ZGZAZGL m ZLZ-3′ 
                 
             
                
                
               
            
           
         
         wherein; 
         each Z is, independently, a modified pyrimidine; 
         each L is independently, for each occurrence, selected from the group consisting of a substituted or unsubstituted C 2 -C 10  linker, a substituted or unsubstituted C 2 -C 8  linker, a substituted or unsubstituted C 2 -C 6  linker, a substituted or unsubstituted C 2 -C 5  linker, a substituted or unsubstituted C 2 -C 4  linker, a substituted or unsubstituted C 3  linker, a G, a C and an A; 
         n is 1 to 5; and 
         m is 1 to 10. 
       
     
     
       18. The aptamer of  claim 17 , wherein each Z is independently selected from 5-(N-benzylcarboxyamide)-2′-deoxyuridine (BndU), 5-(N-benzylcarboxyamide)-2′-O-methyluridine, 5-(N-benzylcarboxyamide)-2′-fluorouridine, 5-(N-phenethylcarboxyamide)-2′-deoxyuridine (PedU), 5-(N-thiophenylmethylcarboxyamide)-2′-deoxyuridine (ThdU), 5-(N-isobutylcarboxyamide)-2′-deoxyuridine (iBudU), 5-(N-isobutylcarboxyamide)-2′-O-methyluridine, 5-(N-isobutylcarboxyamide)-2′-fluorouridine, 5-(N-tryptaminocarboxyamide)-2′-deoxyuridine (TrpdU), 5-(N-tryptaminocarboxyamide)-2′-O-methyluridine, 5-(N-tryptaminocarboxyamide)-2′-fluorouridine, 5-(N-[1-(3-trimethylamonium)propyl]carboxyamide)-2′-deoxyuridine chloride, 5-(N-naphthylmethylcarboxyamide)-2′-deoxyuridine (NapdU), 5-(N-naphthylmethylcarboxyamide)-2′-O-methyluridine, 5-(N-naphthylmethylcarboxyamide)-2′-fluorouridine, and 5-(N-[1-(2,3-dihydroxypropyl)]carboxyamide)-2′-deoxyuridine). 
     
     
       19. The aptamer of  claim 17 , wherein n is 1, 2, 3, or 4; and wherein m is 1, 2, 3, 4, 5, 6, 7, 8, or 9. 
     
     
       20. The aptamer of  claim 17  further comprising at least one, at least two, at least three, at least four, or at least five nucleosides comprise a 2′-OMe. 
     
     
       21. The aptamer of  17  further comprising at least one, at least two, at least three, at least four, or at least five internucleoside linkages are phosphorothioate linkages. 
     
     
       22. The aptamer of  17 , wherein the aptamer binds PDGF with an affinity of less than 10 nM, less than 5 nM, less than 2 nM, or less than 1 nM. 
     
     
       23. The aptamer of  claim 17 , wherein the aptamer inhibits PDGF-mediated phosphorylation of a PDGF receptor. 
     
     
       24. A method comprising administering to a subject in need a therapeutically effective amount of the aptamer of  claim 17 , wherein the method treats and/or prevents a conditions selected from the group consisting of macular degeneration, an ophthalmic condition, fibrosis, a cardiovascular disease and a cancer.